Identification of VP1/2A and 2C as Virulence Genes of Hepatitis A Virus and Demonstration of Genetic Instability of 2C

Emerson, Suzanne U.; Huang, Ying; Nguyen, Hanh; Brockington, A.; Govindarajan, Sugantha; Claire, Marisa St.; Shapiro, Max; Purcell, Robert H.

doi:10.1128/jvi.76.17.8551-8559.2002

Cited by 43 publications

(25 citation statements)

References 21 publications

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“…However, adaptive mutations in this protein highly attenuate the virus for in vivo replication. Such attenuated viruses frequently develop back-mutations or compensating mutations in the 2C protein in vivo and revert to a virulent phenotype (40). Efficient in vitro growth of HAV is also dependent on an adaptive mutation in the 2B protein (2B: A216V) (39).…”

Section: Discussionmentioning

confidence: 99%

Mutations that permit efficient replication of hepatitis C virus RNA in Huh-7 cells prevent productive replication in chimpanzees

Bukh

Pietschmann

Lohmann

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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242

181

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The development of a subgenomic replicon derived from the hepatitis C virus (HCV) strain Con1 enabled the study of viral RNA replication in Huh-7 cells. The level of replication of replicons, as well as full-length Con1 genomes, increased significantly by a combination of two adaptive mutations in NS3 (E1202G and T1280I) and a single mutation in NS5A (S2197P). However, these cell culture-adaptive mutations influenced in vivo infectivity. After intrahepatic transfection of chimpanzees, the wild-type Con1 genome was infectious and produced viral titers similar to those produced by other infectious HCV clones. Repeated independent transfections with RNA transcripts of a Con1 genome containing the three adaptive mutations failed to achieve active HCV infection. Furthermore, although a chimpanzee transfected with RNA transcripts of a Con1 genome with only the NS5A mutation became infected, this mutation was detected only in virus genomes recovered from serum at day 4; viruses recovered at day 7 had a reversion back to the original Con1 sequence. Our study demonstrates that mutations that are adaptive for replication of HCV in cell culture may be highly attenuating in vivo. Hepatitis C virus (HCV) is an enveloped virus with a positivestrand RNA genome (1). Worldwide, more than 100 million people are persistently infected with HCV (2). Infected individuals are at increased risk of developing liver cirrhosis and hepatocellular carcinoma. In a subset of infected individuals it is possible to eliminate the infection by therapy with IFN and ribavirin (3,4). Yet, for the development of vaccines or new therapeutics for HCV, it is of great importance to develop cell culture systems to replicate and propagate HCV. The recent construction of a subgenomic replicon, capable of replication in Huh-7 cells, might represent a first step in achieving this goal (5).The HCV genome (Ϸ9,600 nucleotides) has one ORF, which is flanked by UTRs. Translation produces a polyprotein of Ϸ3,000 amino acids that is cleaved into structural [core (C), viral envelope glycoproteins (E1 and E2)], p7, and nonstructural (NS2-5B) proteins by host or viral proteases (6, 7). The NS2 protein, with the N-terminal third of the NS3 protein, is a protease that mediates NS2͞3 cleavage. The N-terminal part of the NS3 protein, with NS4A as a cofactor, has serine-protease activity and mediates the NS3͞4A, 4A͞4B, 4B͞5A, and 5A͞5B cleavages. The carboxy-terminal part of the NS3 protein contains an NTPase and an RNA-helicase. NS4B is a hydrophobic protein that induces the formation of a cytoplasmic membranous structure where all viral proteins are located and that most likely represents the site of RNA replication (8). The NS5A is a phosphorylated protein believed to be important for viral replication. A short region of the NS5A protein is thought to modulate the host IFN-mediated antiviral response (9). Mutations in this region, the IFN sensitivity-determining region, seems to be associated with the sensitivity of HCV genotype 1b viruses to IFN treatment. The protein ...

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Section: Discussionmentioning

confidence: 99%

Mutations that permit efficient replication of hepatitis C virus RNA in Huh-7 cells prevent productive replication in chimpanzees

Bukh

Pietschmann

Lohmann

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

242

181

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“…These two substitutions were also found for other organs from the same mice (spleen, lung, and heart), suggesting a relevance of these two positions in 2C in the adaptation of FMDV to enable replication in mice. Mutations in 2C have been found upon adaptation of FMDV and other picornaviruses to different environments in cell culture and in vivo (6,8,20,21,28,33,38,46,59). Their possible biological significance is under study.…”

Section: Discussionmentioning

confidence: 99%

Hidden Virulence Determinants in a Viral Quasispecies In Vivo

Sanz-Ramos

Segundo

Escarmı́s

et al. 2008

J Virol

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The characterization of virulence determinants of pathogenic agents is of utmost relevance for the design of disease control strategies. So far, two classes of virulence determinants have been characterized for viral populations: those imprinted in the nucleotide sequence of some specific genomic regions and those that depend on the complexity of the viral population as such. Here we provide evidence of a virulence determinant that depends neither on a genomic sequence nor on detectable differences in population complexity. Foot-andmouth disease virus is lethal for C57BL/6 mice showing the highest viral load in pancreas. Virus isolated from pancreas after one passage in mice showed an attenuated phenotype, with no lethality even at the highest dose tested. By contrast, virus from sera of the same mice displayed a virulence similar to that of the parental wild-type clone and virus isolated from spleen displayed an intermediate phenotype. However, viral populations from pancreas, spleen, and serum showed indistinguishable consensus genomic nucleotide sequences and mutant spectrum complexities, as quantified according to the mutation frequencies of both entire genomic nucleotide sequences of biological clones. The results show that the populations with differing virulences cannot be distinguished either by the consensus sequence or by the average complexity of the mutant spectrum. Differential harvesting of virus generated by cell transfection of RNA from serum and pancreas failed to reveal genetic differences between subpopulations endowed with differing virulences. In addition to providing evidence of hidden virulence determinants, this study underlines the capacity of a clone of an RNA virus to rapidly diversify phenotypically in vivo.

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“…In addition, relatively low concentrations of virus and viral antigen are produced compared to other picornaviruses. Mutations in viral nucleic acid may play a major role in the adaptation of HAV in cell culture (47,76,77,88) and attenuation (79,81). HAV replicates in cell culture without cytopathic signs of infection and without apparent host cell damage.…”

Section: Antigen Detectionmentioning

confidence: 99%

Diagnosis of Hepatitis A Virus Infection: a Molecular Approach

et al. 2006

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SUMMARY Current serologic tests provide the foundation for diagnosis of hepatitis A and hepatitis A virus (HAV) infection. Recent advances in methods to identify and characterize nucleic acid markers of viral infections have provided the foundation for the field of molecular epidemiology and increased our knowledge of the molecular biology and epidemiology of HAV. Although HAV is primarily shed in feces, there is a strong viremic phase during infection which has allowed easy access to virus isolates and the use of molecular markers to determine their genetic relatedness. Molecular epidemiologic studies have provided new information on the types and extent of HAV infection and transmission in the United States. In addition, these new diagnostic methods have provided tools for the rapid detection of food-borne HAV transmission and identification of the potential source of the food contamination.

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Identification of VP1/2A and 2C as Virulence Genes of Hepatitis A Virus and Demonstration of Genetic Instability of 2C

Cited by 43 publications

References 21 publications

Mutations that permit efficient replication of hepatitis C virus RNA in Huh-7 cells prevent productive replication in chimpanzees

Mutations that permit efficient replication of hepatitis C virus RNA in Huh-7 cells prevent productive replication in chimpanzees

Hidden Virulence Determinants in a Viral Quasispecies In Vivo

Diagnosis of Hepatitis A Virus Infection: a Molecular Approach

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